The macrophage-mediated inflammatory response plays a critical role in the development of obesity-related tissue inflammation and insulin resistance. Free-fatty acids (FFAs) are well-characterized factors causing production of inflammatory factors and insulin resistance. Glia maturation factor gamma (GMFG), a member of the ADF/cofilin family of proteins that regulate actin cytoskeleton reorganization, is preferentially expressed in inflammatory cells, but its function in the macrophages immune response remains unclear. In this study, we investigated whether GMFG may affect FFAs-induced inflammatory reaction in macrophages. We show here that silencing of GMFG (80% inhibition of the endogenous expression levels) by transfected with GMFG siRNA significantly enhanced FFAs-induced production of proinflammatory cytokines and chemokines, including TNF-&#945;, IL-6 and IL-8 compared to transfected non-targeting silencing siRNA in human peripheral blood monocytes-derived macrophage as determined by quantitative real time-PCR and confirmed by enzyme-linked immunosorbent assay. These increased inflammatory cytokines resulted from an increased activation of NF-&#954;B and the ERK1/2 MAPK signaling pathway (based on increased NF-&#954;B p65 phosphorylation, I&#954;B&#945;loss and enhanced phosphorylation of ERK1/2 in Western blot analysis) and a reduced phosphorylation of the PI3K/Akt/GSK3-&#946;pathway following FFAs stimulation. Overexpression of GFP-GMFG in GMFG-silenced cells abrogated enhanced phosphorylation of NF-&#954;B p65, ERK1/2 MAPK and reduced phosphorylation of Akt and GSK3&#8722;&#946;. Furthermore, in cells in which GMFG was knockdown, FFAs induced an increase in the expression of SHIP1, a phosphatase that negatively regulates the PI3K signaling pathway, suggesting increased SHIP1 expression may be responsible for the augmentation of inflammatory cytokines following FFAs stimulation. We also show that silencing of GMFG enhances the oxidized low density lipoprotein (oxLDL) induced expression of TNF-&#945;and IL-6 in MDM. Taking together with the data that GMFG is constitutively expressed in macrophages and its expression is down-regulated by FFAs stimulation, GMFG might function as a novel negative regulator through participating in the PI3K/Akt signaling pathway, suggesting that macrophage-specific modulation of GMFG may be beneficial in the treatment of FFAs induced inflammation.